Research Proposal Type 2 diabetes (T2D) is defined by hyperglycemia, caused by peripheral insulin resistance and insufficient release of insulin from the islet ?-cell. Reduced islet ?-cell mass and function have been identified as pivotal events in the progression of insulin resistance toward T2D. Identifying agents regulating islet ?-cell function are critical for developing new diabetes therapies. Our studies focus on the secreted hormone VGF (unrelated to VEGF, and non-acronymic), which we show to control important aspects of islet ?-cell biology-insulin secretion and survival. VGF is expressed in neuroendocrine tissues (hypothalamus, hippocampus), peripheral neurons innervating the gut and adipose, and pancreatic islet cells. Pro-VGF is differentially cleaved by the hormone processing enzymes PC1/3 and PC2; VGF peptides are stored in dense core granules and secreted via the regulated secretory pathway. Targeted deletion of VGF in mouse models has demonstrated VGF regulates energy balance via the CNS; however, the biological significance of VGF expression within pancreatic islets remains unknown. My studies have documented the pharmacological actions of one VGF peptide, TLQP-21, as a novel islet ?-cell secretagogue able to promote the survival of ?-cells in a rodent model of T2D. To assess the physiological function of endogenous islet expressed VGF, we show that siRNA- mediated suppression of VGF in insulinoma cells strongly reduces both fuel and non-fuel stimulated insulin secretion. The central theme of this proposal is to examine the physiological role of islet-derived VGF peptides in regulating insulin secretion from the ?-cell using animal models. The major goals of this proposal are: 1) to fully investigate the mechanism of VGF-mediated regulation of islet ?-cell function; and 2) to examine the impact of loss of a key VGF peptide (TLQP-21) on islet function and survival; and 3) to examine the impact of conditional, ?-cell deletion of VGF on glucose homeostasis in rodent models. In Specific Aim 1, we will determine the mechanism of the insulin secretion deficiency observed upon VGF siRNA suppression utilizing gene expression analyses and ultrastructural examinations. In addition, we will identify which VGF peptide(s) is required for normal islet ?-cell insulin secretory function. In Specific Aim 2, we will use a humanized mouse model in which either the sequence for either full length human VGF or human VGF with a C-terminal truncation lacking the TLQP-21 peptide has replaced the mouse gene at the endogenous locus (knock-in). Studies will be divided between whole animal measurements of glucose homeostasis and isolated islet studies of ?-cell function and survival. In Specific Aim 3, we will generate a novel reagent -a conditional ?-cell specific deletion of VGF using a floxed VGF allele and a tamoxifen inducible, ?-cell specific Cre driver, MIP-Cre/ERT. Using this model we will determine the impact of loss of ?-cell expressed VGF on islet function and glucose homeostasis in a mouse model of insulin resistance. The studies outlined in this proposal will provide novel insight into the biological significance of VGF peptides produced within islet ?-cells. Discoveries into VGF function could translate into a novel treatment strategy for improving glucose control and islet function in T2D. Candidate My long-term vision for my lab is to integrate my expertise in molecular and cell biology with animal physiology to tackle key questions in the field of islet cell biology. My immediate career objective is to obtain a position as a tenure-track assistant professor at a major medical (diabetes) research institution. The studies described in this proposal will provide key training opportunities to afford me a competitive application for faculty position and subsequent funding agencies. My mentor, Dr. Christopher Newgard and his lab have successfully pioneered the use of adenoviral gene delivery tools to study genes and pathways important for the regulation of islet cell function. The Newgard lab also emphasizes the use of rodent models (genetic, diet-induced, and chemically-induced) to study aspects of obesity, insulin resistance, and islet ?-cell dysfunction in the context of whole animal metabolic regulation of glucose homeostasis. The proposal outlined here has taken advantage of many of the Stedman Center's training opportunities, yet continues to move forward into new areas of research for me. Particularly useful, will be my studies using state-of-the-art genetically inducible mouse model to explore the impact of conditional VGF ablation in the islet ?-cell. In this area my prior experience has been limited and Dr. Deborah Muoio (co-mentor) has graciously offered her expertise in training me in this vital area of islet research. Furthermore, as my studies delve deeper into fundamental components of ?-cell function (i.e. insulin secretion) ultrastructural characterization of insulin defects in my model va the Duke Electron Microscopy Core facility will have great impact on my knowledge and skill set as an islet cell biologist. Drs. Newgard and Muoio are fully committed to helping me achieve my long-term goals, allowing me to independently design this proposal and carry this project with me to the next level of an independent faculty position. Thus, through this award, I will gain training and generate crucial reagents necessary to launch my independent research program and obtain a tenure-track assistant professorship.